Many studies have reported the presence of mutations in E2F4 AGC trinucleotide repeats in colorectal cancer bearing microsatellite instability (MSI). The more frequent mutations observed are the deletion or the addition of a trinucleotide AGC and the deletion of 7 trinucleotides (Yoshitaka et al., 1996; Souza et al., 1997; Ikeda et al., 1998; Moriyama et al., 2002). Furthermore, Takashima et al., 2001, studied the impact of E2F4 mutations and observed an increase in nuclear expression, in transcriptional activity as well as in proliferation rate of fibroblasts overexpressing these mutants.

Loss of chromosomal material at 16q22.1 is one of the most frequent genetic aberrations found in breast carcinogenesis suggesting the presence of a tumor suppressor gene (TSG) at this region (Dorion-Bonnet et al., 1995; Iida et al., 1997; Cleton-Jansen et al., 2001). E2F4 is one of the candidate genes localized in this region and therefore was analyzed in different studies. Ho et al., 2001, studied E2F4 protein expression in 10 primary breast carcinomas and 10 metastatic nodal tissues. The authors found a lower E2F4 protein expression in 7/10 primary breast carcinomas and in all (10/10) metastatic nodal tissues when compared to corresponding normal breast tissues. No tumor-specific mutation was detected, but polymorphisms were identified in the polyserine tract of E2F4 (3/11). Ho et al., 2001, further suggested that E2F4 is likely to function as a tumor suppressor in breast cancer. Another group published two complementary and more in-depth studies arguing against the role of E2F4 as a TSG in breast cancer. Indeed, Rakha et al., 2005, used a Multiplex Amplifiable Probe Hybridization (MAPH) method to measure DNA copy-number at chromosome arm 16q22.1 in forty-nine invasive lobular, low-grade invasive ductal or tubular breast carcinoma samples. No correlation was detected between the expression of E2F4 with its gene's copy number. Likewise, no significant loss or decrease in E2F4 protein levels was observed in malignant tissues. However, the authors did describe a correlation between increased nuclear expression of E2F4 and tumors with higher histological grade and positive lymph node disease whereas E2F4 was expressed in both the nuclei and cytoplasm in normal mammary epithelial cells, thus suggesting an oncogenic rather than a tumor suppressor role for this factor in breast cancer. The same group (Rakha et al., 2004) also analyzed 265 breast carcinomas for E2F4 protein expression and found a correlation between increased nuclear expression of E2F4 and indicators of poor prognosis including larger tumor size, grade 3 lesions, lymph node stage and poorer Nottingham prognostic index group. Increased E2F4 expression was also seen in association with the development of recurrent disease, distant metastasis and poorer outcome including poorer overall survival time and shorter disease-free interval.

Using serial analysis of gene expression (SAGE), Waghray et al., 2001, found that E2F4 was overexpressed by more than five-fold in prostate tumor tissues compared to the normal surrounding tissues. Immunohistochemistry analysis further revealed strong E2F4 staining in epithelial cells of tumor glands as opposed to weak to no staining in normal glands.